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COVID-19

A primer into the literature

To Our Students in the Department of Biology

March 25 2020, Version 1.0

@biologyatCU

Special Presentation, March 21, 2020

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IMPORTANT TO READ BEFORE PROCEEDING

Many people made very substantial contributions to this document (see Acknowledgement slide at end).

They made these contributions in good faith in an academic spirit.

I however (Mike Hallett) take full responsibility for all errors, omissions and misstatements of any kind.

I ask that you read the next slide very carefully before going through the rest of the presentation.

If you would like to send comments and corrections, or build new versions, you can reach me on the Department of Biology’s slack workshop or by email michael.hallett@concordia.ca

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IMPORTANT TO READ BEFORE PROCEEDING

Our Goals Here

A disease and how it affects society is multifaceted (and therein lies the problem).

Our goal here is not to give definitive answers, advocate for a specific policy, suggest clinical treatment or influence decision making by our government, university, medical system, or other involved parties.

We are not ourselves experts on any topic in this document. We have knowledge of the fundamental biology and scientific methods. We feel that this information will help you better explore for yourself this disease as it progresses in our communities.

Our goal here is not to make you “arm chair” epidemiologists, drug designers, clinicians or politicians. You came to Concordia to study biology, and these concepts underlie many of the facets of a pandemic.

Our goal here is to direct you towards researchers and specific research efforts that have informed on SARS-CoV-2-2019, COVID-19. This is intended as a primer into the literature; not as a self-contained lecture about any one concept.

Our understanding is fluid and changing rapidly. This is not a complete list of resources, nor is it intended to be. We cannot hope to keep up. We are just providing starting points into the literature.

The science is moving very fast but some of these studies are very small and speculative, often just case reports.

Many are from reputable labs with reputable leaders and reasonable results, but they may not have been thoroughly reviewed (or reviewed at all) by usual academic and medical standards.

Vaccines and therapeutics take a long time to develop. The intention of the research community currently is to make these “preprints” available quickly in venues such as medRxiv and bioRxiv, so that the research community can move with agility forward. Therefore we need to be critical of what we read.

In this document, we tried to avoid citing preprints and non-refereed material. This document is not refereed and so it too should be read with caution. We sincerely hope to assist your journey to explore the biology of covid-19.

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Maybe it would help if we phrase this an exam :-) Old habits die hard.

Open Book (MOCK) Exam Question #1 [33 marks out of 100], 18 months to complete.

Throughout this document, we have made a concerted effort to cite and link to papers from the primary scientific literature with a preference for journals that are regarded as “prestigious” and to have “high impact”.

In some cases, we reference manuscripts that appear in preprint servers such as bioRxiv, arXiv, and medRxiv.

In some cases, we link to the general media, specifically articles accessible to lay-people. We have tried to choose those articles that supply links back to the primary scientific literature.

There are several examples in the presentation where no link back to the scientific literature is provided.

Find 5 examples of references to manuscripts in this presentation that have not been refereed. Find 5 examples of papers that, although they have been refereed and appear in journals, nevertheless appear to have very little data supporting their findings.

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Technical Note

This document provides many links to papers in the scientific literature.

Although many journals now offer open access, some remain behind a

paywall.

Whenever possible, we have chosen manuscripts that are open access.

In other cases, the university has subscriptions to many of the journals

that are not open access.

To access papers in these journals, you would need to use VPN to access the Concordia internet or login to the library with your Concordia Netname.

VPN is straightforward and IITS offers instructions here.

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Outline

  1. Definition of covid-19, corona virus disease, SARS-CoV- 2
  2. Molecular mechanisms
  3. Epidemiology
  4. Molecular tests for detection
  5. Drugs and therapeutics
  6. Clinical presentation of covid-19
  7. Effects of pandemic on the environment

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  1. Definitions and Basic Facts

covid-19, corona virus disease, SARS-CoV- 2

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  1. covid-19 vs coronavirus disease vs SARS-CoV-2

Disease: coronavirus disease (COVID-19)

Virus: severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)

The World Health Organization (WHO) is an excellent source of information that has been packaged for general audiences. It provides much more information about the history of covid-19, SARS-CoV-2.

SARS is caused by a coronavirus called SARS-associated coronavirus (SARS-CoV). SARS was first reported in Asia 2003.

SARS spread to more than two dozen countries in North America, South America, Europe, and Asia before it was contained. WIth a mortality rate near 10%, it killed 774 people including 44 Canadians.

Since 2004, no cases of SARS have been reported.

The Center for Disease Control and Prevention is another excellent website with excellent science packaged for the general public.

All coronaviruses so far have been linked to respiratory diseases, from the common cold to pneumonia (Graham, 2013 Nature Rev Microbiol)

.

@WHO @GovCanHealth

@CDCgov

We recommend:

@sante_qc

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SARS-CoV-2 2019 - A primer for #COVID-19

Why do we recommend this report as a good starting point, especially for non-specialists?

The explanation is very nice, easy to understand. Suitable for your family and friends, but also -

All of the claims made in the article (to the best of our knowledge) are backed up by citations into the primary scientific literature.

The citations themselves are to papers that appear (to the best of our knowledge) in reputable journals.

The senior authors of these papers are well known in their field.

The explanation is comprehensive, touching on many aspects of the disease.

That does not mean that everything in this report is necessarily true. Because of the urgent situation, findings are being released with less rigorous testing than usual. Many statements are therefore very tentative and can change within days as new data arrives.

We can question but defer to experts.

There are many other critical questions you must ask whenever you read any paper or report, including this presentation. Eyes wide open.

1. What do we know about the coronavirus?

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1. What do we know about the coronavirus?

Enveloped, positive (+) strand RNA virus. There are 7 coronaviruses known to infect people, 4 only cause mild disease similar to the common cold (rhinoviruses), but 3 are lethal: SARS, Middle Eastern Respiratory (MERS), and SARS-CoV-2.

This is an excellent resource from The Economist for lay-people (your friends and family) that explains the basics of the coronavirus.

Coronaviruses cause respiratory diseases from colds to pneumonia (Graham, 2013 Nature Rev Microbiol) [Technical]

A classic reference for this field is: Lai MM, Cavanagh D (1997). "The molecular biology of coronaviruses" [Technical]

[CDC, 2020]

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SARS-CoV-2 is highly infectious compared to other coronaviruses.

  • SARS-CoV-2 virus can be found actively replicating much earlier than its “cousin” SARS-CoV (day 5 vs. 7-10) (Pan 2020 The Lancet Infect Dis, Peiris 2003 Lancet).
    • This is cause of concern about contagion/virus spreading from asymptomatic individuals.
  • ~1000 times more SARS-CoV-2 viral particles than SARS-CoV were found in a single swab.
  • Unclear for how long patients can “shed” viruses after symptom resolution.
  • SARS-CoV-2 appears to be generally less lethal than either SARS-CoV (10% death rate) or MERS (34 %)
  • Interesting interview with virologist David Ho for all audiences from Caltech.

[D. Ho is a world expert in HIV/AIDS and viral epidemics at Columbia U and a Caltech Trustee.]

1. What do we know about the coronavirus?

Electron micrograph of SARS-CoV inside an infected cell. Source: C.S. Goldsmith, CDC

Colorized scanning electron micrograph of SARS-CoV-2 (yellow). Viral particles exiting from cultured cells (blue/mauve). Source: NIH/NIAID.

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Zoonotic spillover: animal-human disease transmission

The majority (94%) of zoonotic viruses described to date (n = 162) are RNA viruses, which is 28 times higher than the proportion of RNA viruses among all vertebrate viruses recognized

Human-to-human transmission was observed for 20% of zoonotic viruses. Many zoonotic diseases are self-limited, i.e. there is no human-to-human transmission. e.g. rabies.

The most frequent zoonotic spillovers are from rodents, primates and bats. Bats harbour an unusual number of zoonotic viruses; the reason is not clear, but one hypothesis suggests that the bat immune system has evolved to withstand the high body temperatures that occur during flight.

Most human zoonotic viruses are reported from at least two different orders of mammals. Domestic animals are frequent intermediate hosts between wild reservoir species and humans.

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For further reading, we recommend:

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Zoonotic disease spillover I

Zoonotic spillover is usually linked to human activity and habitat destruction: stressed animals are more likely to express the virus, and animals whose habitat has been destroyed or profoundly modified are more likely to come in contact with people. Zoonotic spillovers happen in regions with rapid habitat destruction.

There have been many minor zoonotic disease spillovers with limited human-to-human transmission, e.g. the Nipah virus in India: upsurges 2001 (45 deaths), 2007 (5 deaths), 2018 (17 deaths, from 2 bat-human contacts)

Viruses with high host plasticity (i.e. that accept many hosts) are more likely to be transmitted human-to-human.

We recommend: [Allen et al. (2017) Nature Communications]

Here are two reports accessible to lay-people that we recommend:

[The Guardian (March 18, 2020) “Tip of the Iceberg”]

[Ensia (March 18, 2020) “Destruction of habitat and loss of diversity…”]

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1. What do we know about the coronavirus?

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Zoonotic disease spillover II

The 2002 SARS epidemic brought Coronaviruses under the spotlight: it was the first time a dangerous coronavirus emerged in human populations.

The initial human cases of SARS were caught from civet cats. Later, antibodies against SARS were found in three horseshoe bat species suggesting that bats are the natural reservoir species, which cross-infected the civets. FInally in 2013, a coronavirus 97% identical to SARS was found in bats, confirming their role as natural hosts.

There are hundreds of bat-infecting corona viruses, some can cross into other animals, including pigs, and cause veterinary disease.

SARS-CoV-2 is 96% identical to a coronavirus identified from horseshoe bats, which implies that bats are likely the natural reservoir for the virus, even if it might have passed through an intermediate host (perhaps pangolins in which SARS-CoV-2 like viruses have also been identified) prior to infecting humans.

We recommend the following article accessible to lay-people from Scientific American.

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Bat-borne viruses

Covid-19 is the sixth bat-borne virus to cause recent outbreaks of human disease:

  • Hendra (1994)
  • Nipah (1998, 2001, 2007, 2018)
  • SARS (2002) a coronavirus, intermediate host: civet cats
  • MERS (2012) a coronavirus, intermediate host: dromedary camels
  • Ebola (multiple reinfections, some leading to outbreaks, since discovery in 1976, most recent in 2018)

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Clinical manifestations

Most patients show Influenza-like illness (ILI) and the most commonly reported symptoms include:

  • Fever
  • Fatigue
  • Dry cough
  • Myalgias (muscle pain)
  • Dyspnea (Difficulty breathing)
  • Sore throat

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1. What do we know about the coronavirus?

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Serious manifestations and complications include:

1. What do we know about the coronavirus?

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Tissue Morphology

Observations during Jan-Feb 2020 indicated that the greatest severity of lung disease manifested ~10 days after initial onset of symptoms with pneumonia hitting both lungs

[Pan et al. (Feb 13, 2020) Radiology]

Lungs are progressively damaged and eventually, their extensive scarring leads to the need for mechanical ventilation (Namendys-Silva, 2020 The Lancet)

Image of the SARS-CoV-2 viruses (blue) from the first US COVID-19 case

(Source: New York Times)

1. What do we know about the coronavirus?

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Why are some COVID-19 cases so much worse than others?

We do not have a precise answer yet.

However, once the virus has attached itself to the lungs and begins to damage the inner alveoli, immune cells are attracted to the damaged site.

In most cases, the immune system would be able to eventually stem infection and lung damage would be eventually repaired.

However, in some individuals, abnormal release of immune mediators, called a cytokine storm, is causing damaging inflammation, pneumonia and shortage of breath.

A nice article for all audiences outlining this process was published in The Scientist.

[source: The Scientist]

1. What do we know about the coronavirus?

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Since SARS-CoV-2 is evolving, there can be no static facts.

As its genotype changes, its phenotypes may also change.

As it moves into new geographical and ecological niches and interacts with different human populations, key properties

of the virus will change.

On the next page, we show a movie from Bedford and his colleagues created via their beautiful website

nextrain.org/ncov to follow SARS-CoV-2 evolution across time and space.

This group sequenced the (RNA) genome of several hundred covid-19 positive patients (on going).

They then used bioinformatics techniques to build a molecular phylogeny (a branching tree).

You can watch the nucleotide or amino acids change across the genome, over time, across geography.

This evolution makes it very difficult to pin down global “timeless” properties eg it might have a certain mortality rate X in China but a mortality rate Y in Europe.

This analysis can provide pseudo-real time insight into how the virus is changing at the molecular level, and

provide means to estimate via mathematical models “undocumented” (hidden) community transmission of

the disease (see this example if you are interested for Washington State, USA).

Peter Thielen’s analysis (John Hopkins) suggests the SARS-CoV-2 is not evolving very fast, which is good news for

the feasibility of a vaccine [Washington Post (March 24 2020)].

1. What do we know about the coronavirus?

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1. What do we know about the coronavirus?

How long will SARS-CoV-2 survive on surfaces?

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1. What do we know about the coronavirus?

So your neighbour asks you,

“What about dogs? Can they be infected? Can they carry the virus?”

How do you answer?

In some cases, the science does not yet exist to answer a question definitively.

Any answer should reflect that fact, and our scientific uncertainty, very clearly.

We recommend:

The short answer is that there is little evidence that dogs can be infected by or carry the virus currently. However, we know the virus can jump between host species.

However, from the previous slide, the virus can survive on several surfaces for a considerable length of time.

So a dog that is petted or interacts with other dogs could potentially be a carrier, similar to their potential of spreading poison ivy oils. At time of writing, the community is uncertain.

How do you answer your neighbour?

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2. Molecular Mechanisms

Before we begin, we recommend the following Twitter handles to follow.

They are the very best scientific journals and they regularly put out great simple

summaries of new results.

@nature

@ScienceMagazine

@CellPressNews

@JAMA_current

Basic Research

Clinical

Research

@NEJM

For a very good comprehensive view, we would recommend @EricTopol

We will let you decide if you trust this physician scientist, author and editor

And of course PubMed at the NCBI

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What do we know about how SARS-CoV-2?

Here is its RNA-based genome. Stop to think of what just 31,000 bp of RNA and 15 genes have already managed to do in < 1 year. By comparison, the human genome is ~3.6 billion bp with ~20,000 genes.

2. Molecular Mechanisms

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2. Molecular Mechanisms - Genomics

[Note: if you have a friend who favours “certain” conspiracy theories,

you might want to sit (distantly) with them and explain this paper to them.]

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How does it get into cells and what does it do?

2. Molecular Mechanisms

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  • The SARS-CoV-2 virus may have acquired the capacity to infect many cell types also through mechanisms independent of the

Angiotensin-Converting Enzyme 2 ACE2 receptor.

  • If the spike proteins from coronaviruses interact with several factors (Park 2019 Nature Struct Mol Biol, Zhou 2020 Nature) that would provide alternate entry routes into cells.

  • Indeed, proteolytic cleavage of the spike protein increased coronavirus infectiousness by promoting fusion with the cell membrane and cell entry (Belouzard 2019 PNAS).

  • In the SARS-CoV-2 virus, the spike protein contains a novel cleavage site for furin.
    • This would allow infection of all furin-expressing cells, independently of the expression of the ACE2 receptor.
    • The key is that… furin is expressed by most cells! This hypothesis must be further explored.

  • (Below we return to protease inhibitors as a therapeutic.)

2. How does this work at the molecular level?

[Qing 2020 Trends Immunol a clear brief summary of the new developments on proteolytic cleavage. For expert audience.]

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In SARS-CoV pathology there seem to be a clear component of pathological immunity combining multiple effects:

Overactivated innate immunity → cytokine storm → tissue damage

AND

Depressed T-cell response → compromised anti-viral response +

lack of control of cytokine storm

AND

Potential for long term lung damage initiated by certain antibody responses →

important for long-term organ damage

The next 5 slides will go into more detail and require an interest for the pathological mechanisms.

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2. SARS-CoV immunology: inflammasomes and cytokine storms

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Inflammation is both anti-viral (attracts immune cells to the site of infection to attack the virus) and pro-viral (induces tissue damage facilitating viral exit and spread).

  • Infection of nearby motile cells like macrophages and dendritic cells (attracted by the infection) can carry the virus to distant body parts and potentially spread infection.
  • Immune cells, e.g., circulating lymphocytes, monocytes, and macrophages have been found to contain SARS-CoV (Gu 2005 J Exp Med). Their capacity to spread disease needs to be determined.

This is what normally happens in a viral respiratory tract infection:

  • Macrophages are activated. Pro-inflammatory macrophages amplify the response. Anti-inflammatory macrophages promote wound healing via synthesis of matrix metalloproteinases, growth factors and TGF-beta. As the response continues, pro-inflammatory macrophages increase until the virus is eliminated. Eventually, they produce anti-inflammatory interleukin-10 (IL-10) to restore homeostasis.

  • At the same time, innate dendritic cells phagocytose the virus and migrate to the closest lymph nodes, process the antigens into peptides and present them to circulating T-cells. Peptide-specific T-cells are activated, migrate to the infection site, where they produce cytokines interferon-gamma (inhibits viral replication, enhances antigen presentation), TNF-alpha, IL-2, chemokines CXCL-9-11 (recruit more innate and adaptive cells), and cytotoxins perforin and granzyme (lyse infected cells). The more T-cells are specific for the virus, the faster the clearance.

  • At the same time, B-cells are activated and eventually produce neutralizing antibodies.

  • Memory B- and T-cells are formed (will block future infections from the same or similar pathogens).

2. SARS-CoV immunology: background

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2. SARS-CoV immunology: The bases of the cytokine storm

SARS-CoV patients display severe inflammation with the markers below, but the onset mechanism is not known.

  • Higher levels of proinflammatory tumor necrosis factor α (TNF‐alpha), CXCL‐10, IL‐6, and IL‐8.
  • Reduced anti-inflammatory IL-10 (Kong 2009 Virus Res).
  • In severe cases IL‐1, IL‐6, IL‐12, Interferon gamma, and transforming growth factor‐beta) and chemokines (CCL2, CXCL9, CXCL10, and IL‐8) were further elevated (Chien 2006 Respirology, Zhang 2004 Infectious Immun).
    • IL-1 is linked to inflammasome activation.

  • Delayed anti-viral interferon response (Menachery 2014 mBio, Channappanavar 2016 Cell Host Microbe).
  • Reduced CD4+ and CD8+ T-cell response ((Li 2004 J Infect Diseases, Li 2003 Chinese Med J).
    • partly due to inefficient dendritic cell migration (due to high levels of virus-induced prostaglandin 2).

Similar patient presentation is seen in SARS-CoV-2 infection (Liu 2020 J Med Virol, Huang 2020 Lancet). Consider that individual response varies due to unknown factors, potentially linked to genetics, health condition, age, life style (e.g. smoker) and history.

Tisoncik 2012 MMBR, 76(1), 16–32.

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2. The inflammasome link to cytokine storms

The SARS-CoV-2 virus contains two viroporin proteins called E (envelope) and ORF3a needed to release viruses from infected cells. They are highly proinflammatory ion channels that activate the NLRP3 inflammasome.

SARS-CoV-2

E viroporin

In particular, ORF3a is able to induce the two signals needed to activate the NLRP3 inflammasome (FASEB J 2019)

  • pro-interleukin 1 beta synthesis
  • its cleavage to mature interleukin 1 beta by the caspase 1 enzyme.

SARS-CoV-2 ORF3a

ORF3a also activates NF-kB, Golgi fragmentation, increases intracellular vesicles, ER stress and apoptotic cell death (Virus Research 2008, PLoS One 2009).

The pro-inflammatory signals from the NLRP3 inflammasome, especially with low T-cell activity, can escalate to a cytokine storm that severely damages the lung tissue (acute respiratory distress syndrome, ARDS, followed by Acute lung injury, ALI) and requires artificial ventilation [The Lancet (Feb 27 2020].

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2. What about the anti-virus antibodies of the adaptive response?

Perhaps counter-intuitively, antibodies against the viral spike protein (S) may contribute to acute lung injury, instead of targeting the virus for elimination.

Anti-spike antibodies (S-IgG) block the virus, and at the same time induce a sub-class of pro-inflammatory macrophages that damage lung tissue at the expense of the wound-healing macrophages. The mechanism is unknown (Liu 2019 J Clin Invest).

Although unproven, this phenomenon may explain the worse prognosis of older individuals and higher severity of SARS in China (not necessarily in Italy): individuals with previous encounters with similar viruses may have the capability to produce pro-inflammatory antibodies. This would be an instance where immunological memory may not be useful.

The specificity of such inflammatory antibodies may be restricted to certain portions of the spike protein. Indeed, neutralizing antibodies against the receptor binding portion of the spike protein yielded long term protection in animal models, but their long-term effects on the lung is unclear (Du 2007 Vaccine).

Pathological changes of the lung tissue from S-IgG. Symptoms of acute diffuse alveolar damage. From (Liu 2019 J Clin Invest)

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2. The last step: decreased immunological memory

Normally, encounter with a pathogen leaves behind memory T-cells that can be activated upon pathogen re-encounter. In this instance, virus-specific T-cells rapidly produce interferon-gamma and CXCL9-11 chemokines that attract innate and peripheral T-cells.

SARS-CoV patients develop specific memory T-cells lasting at least four years. However, memory B-cells were found to decrease over time (Chen 2005 J Immunol).

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3. Epidemiology of SARS-Cov-2

Before we begin, let us recommend the following organizations and people to follow.

Firstly, Dept of Biology’s very own @VDumeaux is an epidemiologist. She’d be happy to answer your questions.

The Dalla Lana School of Public Health @UofT_disph has is an excellent source

for Canadian public health/epidemiology related to covid-19.

In particular, @DFisman . He was very vocal about implementing social distancing in Canada.

Another good person in Canada to watch is Andre Picard @picardonhealth

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3. Epidemiology of SARS-Cov-2

The current approach to control the pandemic is perhaps broadly based on two components:

non-pharmaceutical interventions + disease diagnostics

(eg social distancing) (eg throat swab-based RNA testing)

This “flattens the curve” and “buys us time” to develop

pharmaceutical interventions + herd immunity

(eg a vaccine) [Fine et al (2007) Clinical Infectious Diseases]

[Al Jazeera (March 20, 2020)]

This section links to efforts in the scientific community related to these concepts.

An important concept

to learn

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3. First off, what is Epidemiology?

A good place to start is to learn a bit about the basic concepts of how diseases spread in the population. While you are reading, imagine answering the following exam question (in a crowded arena filled with desks with actual people beside you).

Open Book Mock Exam Question #2 [33 marks out of 100], 18 months to complete.

In one million words or less, explain why measuring infection rates in a population can be difficult? Explain why measuring the mortality rates in a population is very difficult. Explain why risk factors are difficult to measure?

Make sure that your answer thoroughly covers the three central concepts of pandemics

  1. transmission rate (R0; the number of newly infected people from a single case)
  2. the case fatality rate (CFR; the percent of cases that result in death) and
  3. how to determine whether asymptomatic transmission is possible.

You may make use of resources such as this primer designed for lay-people [The Economist] and more technical descriptions here.

Also, we recommend Epidemiology by Leon Gordis [a textbook covering the basics] and

Principles of Epidemiology in Public Health Practice [A textbook recommended by the CDC].

Practice Social Exam Distancing - that is - NO CHEATING.

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3. Epidemiology of SARS-CoV-2

The current approach to control the pandemic perhaps broadly based on two components:

non-pharmaceutical interventions + disease diagnostics

(eg social distancing) (eg throat swab-based RNA testing)

This “flattens the curve” and “buys us time” to develop

pharmaceutical interventions + herd immunity

(eg a vaccine)

let’s start here

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A classic paper: Nonpharmaceutical Interventions Implemented by US Cities During the 1918-1919 Influenza Pandemic Markel et al. JAMA (2007)

Context A critical question in pandemic influenza planning is the role nonpharmaceutical interventions might play in delaying the temporal effects of a pandemic, reducing the overall and peak attack rate, and reducing the number of cumulative deaths.”

Results … [text removed] The cities that

implemented nonpharmaceutical interventions

earlier had greater delays in reaching peak

mortality (Spearman r=-0.74, P < .001), lower

peak mortality rates (Spearman r=0.31, P=.02),

and lower total mortality (Spearman r=0.37,

P=.008).”...

Conclusions …[text removed] “The historical

record demonstrates that when faced with a

devasting pandemic, many nations, communities,

and individuals adopt what they perceive to be

effective social distancing measures or

nonpharmaceutical interventions …”

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Some similarities with the influenza pandemic or why is it so difficult to prevent respiratory infections from spreading?

Soper GA. The lessons of the pandemic (Science 1918)

  • “The great complexity and range in severity of the respiratory infections confuse and hide the danger.”

  • “The waste products of influenza containing the infective virus are not deposited in a vessel or sewerage system where they can be properly dealt with as in typhoid. The excreta of the nose and throat are projected into the air and allowed to pollute the hands, the food, the clothing and, in fact, the entire environment of the infected person. This is done unconsciously, invisibly, unsuspectingly.”

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Emergency hospital during influenza epidemic, Camp Funston, Kansas. Otis Historical Archives, National Museum of Health and Medicine.

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Non-pharmaceutical intervention. For example, social distancing.

The next 3 slides look at a paper that has perhaps been influential w.r.t. government policy to address the pandemic.

“... apply a previously published microsimulation model to two countries: the UK (Great Britain specifically) and the US”

“Two fundamental strategies are possible: (a) mitigation, which focuses on slowing but not necessarily stopping epidemic spread … (b) suppression, which aims to reverse epidemic growth …

3. Epidemiology - the effect of non-pharmaceutical interventions

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Unmitigated

Mitigated

Ferguson et al. (March 16, 2020) (Continued)

3. Epidemiology - the effect of non-pharmaceutical interventions

Basically, if we do nothing, everyone gets sick at once and a lot die.

But this is only part of the problem (and where we are right now)....

Next slide please….

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If we clamp down with interventions, the infection rate goes down (hospitals catch up).

But the pathogen is still in the environment and there are still carriers.

If we release, interventions rise (hospitals fill up).

It’s a very chaotic system. Here the authors are showing how ICUs will fill across time under

different strategies.

Ferguson et al. (March 16, 2020) (Continued)

3. Epidemiology - the effect of non-pharmaceutical interventions

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Ferguson et al. (March 16, 2020) (Continued)

3. Epidemiology - the effect of non-pharmaceutical interventions

So here the authors are

showing how the number

of ICU cases goes up and

down in response to

clamping down with non-

pharmaceutical interventions

and then releasing the interventions.

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So one plan is to clamp-release repeatedly.

There are ~11 million people in Hubei, China [wikipedia]. There are currently less than 100K documented infections [John Hopkins map, March 25].

So when interventions like social distancing are relaxed early on, the vast majority of the population remain non-infected individuals (11 million minus 100 thousand). So the clamp-release

cycles would have to be tightly controlled.

Ferguson et al. (March 16, 2020) (Continued)

3. Epidemiology - the effect of non-pharmaceutical interventions

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It might take quite a while to achieve herd immunity [Fine et al (2007) Clinical Infectious Diseases].

If that is the case, the virus will have time to evolve. If so, previously infected may not remain

immune. Early indications is that it is not evolving particularly fast. Here are two very recent

reports on this subject: [Washington Post (March 24 2020)] [Trevor Bedford (March 25) 2020].

There can be other complications. See for example the slide 33 marked with

Molecular Mechanisms 2. What about the anti-virus antibodies of the adaptive response?

Ferguson et al. (March 16, 2020) (Continued)

3. Epidemiology - the effect of non-pharmaceutical interventions

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3. Epidemiology of SARS-Cov-2

The current approach to control the pandemic perhaps broadly based on two components:

non-pharmaceutical interventions + disease diagnostics

(eg social distancing) (eg throat swab-based RNA testing)

This “flattens the curve” and “buys us time” to develop

pharmaceutical interventions + herd immunity

(eg a vaccine)

this next topic

needs no introduction

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3. Epidemiology - the effect of non-pharmaceutical interventions

“Flattening the curve”

An accessible animation illustrating how disease spreads with and without containment.

It changes the patterns every time you run it: Washington Post

grey: susceptible people

orange: sick people

pink: recovered (immune?) people

Source: Washington Post

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3. Epidemiology - the effect of non-pharmaceutical interventions

In addition to social distancing, there are other non-pharmaceutical interventions.

For example, we would protect with greater rigour individuals

  1. at elevated risk of contracting the disease
  2. at elevated risk for severe forms of the disease.

Of course, we need to learn “on the fly” what these risk factors are and how to protect.

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Data on mortality rate by age group shows that risk increases with age.

Hubei, China

[The Economist, March 12]

It is obviously difficult to test for risk factors associated with covid-19, since manipulative experiments can't be done. So, instead, one looks at correlation of various underlying conditions in patients vs the general population.

These epidemiological studies show that fatalities showed a higher than expected association with hypertension, diabetes, chronic lung disease and cancer, suggesting that these conditions could increase risk of serious disease.

Mortality rates appear to be higher for men than for women and might be higher for smokers.

3. Epidemiology - Risk factors for severity of the disease

We recommend this article for further reading [Yang et al (March 12, 2020) IJID].

USA

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3. Epidemiology - Risk factors for severity of the disease

The data is always changing, the virus evolves, and it infects different populations, with different

genotypes, different lifestyles/socioeconomic factors in different environments. This is problematic,

because clinicians still need to make decisions based on very little information.

And then these factors can in turn interact with sex, age, BMI and other variables to create a giant

interconnected network of influences.

For example, in China, there remains a considerable amount of smoking [Li et al. (2011) NEJM]. In 2010,

52.9% men smoked vs. 2.4% women.

Could this explain why there is some evidence of the cigarette smoking might affect the expression

patterns of ACE2 [Wang et al. (2020) PrePrint]

Are covid-19 patients, who are also hypertensive or diabetic and treated with ACE2-stimulating drugs

at increased risk of severe disease? [Fang et al. (March 11 2020) The Lancet]

Ibuprofen as a risk factor ?!? [The WHO March 18 2020]

So if you are diabetic, or asthmatic, what should you do? What if you have hypertension and are taking ACE inhibitors, should you continue with your medication? (Hypertension Canada says yes, other sources raise the question - look for them).

exam hint

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Are men more affected than women?

Possibly. Data is still limited, but it seems that while man and women are equally infected, men may die more of COVID-19 than women. This is consistent with other SARS and MERS coronaviruses.

Also, ACE2 gene is on the X chromosome, making men potentially more susceptible.

3. Epidemiology - Risk factors for contracting the disease

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3. Epidemiology - Risk factors for severity of the disease

[Backhaus (March 13 2020) Medium]

This is an interesting article looking at deaths in Italy versus South Korea.

Intending no disrespect to the author, it would be prudent to ask yourself while reading it

  • Has this work been refereed and reviewed by experts?
  • What are the qualifications of the author?
  • Where is the data he is analyzing coming from?
  • Is there a Methods section describing his statistics and processing of the data?

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3. Epidemiology - Will warm weather slow or ablate its spread?

The next few slides look at a review paper that examines how weather impacts the

spread of certain infectious diseases. The review provides insight into the molecular mechanisms that exist primarily at the interface between the pathogen, our epithlial lung cells and our immune system.

This is for the northern hemisphere. SAR-CoV-2 indicated by red arrow.

Does this mean it will slow down (eg infection rate drops)? Can it “disappear”? Will it come back next winter? This manuscript provides some molecular level insight that perhaps underlie answers to these questions.

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3. Epidemiology - Will warm weather slow or ablate its spread?

Environmental factors, including temperature and humidity, modulate host intrinsic, innate, and adaptive immune responses to viral infections in the respiratory tract.

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3. Epidemiology - Will warm weather slow or ablate its spread?

So there is a molecular basis to understand how weather will affect the ability of covid-19 to infect individuals.

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3. Epidemiology - tracking the disease in the population

Johns Hopkins Coronavirus Resource Center

The data science surrounding the rate of infection in warm versus cool climates will be interesting. covid-19 is expanding at the equator and southern hemisphere with strong economic effects.

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3. Epidemiology - tracking the disease in the population

The WHO’s recommended method

Ok ok you should watch this too.

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3. Epidemiology of SARS-Cov-2

Recall our aims in this section:

non-pharmaceutical interventions + disease diagnostics

(eg social distancing) (eg throat swab-based RNA testing)

This “flattens the curve” and “buys us time” to develop

pharmaceutical interventions + herd immunity

(eg a vaccine) [Fine et al (2007) Clinical Infectious Diseases]

[Al Jazeera (March 20, 2020)]

This section links to efforts in the scientific community related to these concepts.

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3. Epidemiology - tracking the disease in the population

Other Bioinformatic Resources

Worldometer Live Coronavirus Tracker: https://www.worldometers.info/coronavirus/

  • Run by an international team of developers, researchers, and volunteers with the goal of making world statistics available

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3. Epidemiology - tracking the disease in the population

Bioinformatic Resources

  • Up to date epidemiology for Canada, province by province: here
  • Up to date visual world map report by the Center for System Science and Engineering: here.

Distribution of confirmed and probable cases of COVID-19 by province or territory in Canada as of March 21, 2020, 9:00 am EST

Source: Government of Canada

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3. Epidemiology - tracking the disease in the population

Bioinformatic Resources

COVID-19 Canadian Outbreak Tracker: https://virihealth.com/

  • Compiles information from Official Government Health Ministry websites on a daily basis
  • Associated map

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3. Epidemiology - tracking the disease in the population

Bioinformatic Resources

China National Center for Bioinformation:

  • Features comprehensive integration of genomic and proteomic sequences as well as their metadata information from the GISAID, NCBI, NMDC and CNCB/NGDC.

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3. Epidemiology of SARS-CoV-2

The current approach to control the pandemic perhaps broadly based on two components:

non-pharmaceutical interventions + disease diagnostics

(eg social distancing) (eg throat swab-based RNA testing)

This “flattens the curve” and “buys us time” to develop

pharmaceutical interventions + herd immunity

(eg a vaccine)

let’s work towards an understanding of this now

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3. Why is testing important?

Testing means applying an assay (“non-molecular” eg fever or “molecular” eg measure RNA in a throat swab) to determine if any individual is infected.

Generally the more testing is done, the sooner infected individuals can be quarantined and classic epidemiology methods can be used to track down everyone the person has come into contact with.

Testing also provides important statistics for tracking the disease and estimating mortality rates (a very difficult estimation problem).

We recommend to start with this article [Subbaram (March 23, 202), Nature]. The next few slides

explore why testing is important and tie it back to the epidemiology of the disease.

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4. First, “non-molecular” tests for detection

A reliance on self-diagnosis (eg you realize you have a dry cough and fever) for detecting disease

allows the virus to spread. Here is an antidote close to home.

Among many other issues, it is often not clear how long it takes for different symptoms to manifest themselves.

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4. Testing allows us to measure the infection rate

There are many questions about whether asymptomatic people can spread the virus and how long one remains contagious.

A recent manuscript uses data from our colleagues in China that suggests a very high frequency of SARS-CoV-2 infections may be present in the population: [Li et al. (March 16, 2020) Science]

So this might suggest that non-molecular tests are insufficient and molecular tests need to be expanded.

Note the very advanced mathematics that are used (see Methods section).

R0 (basic reproduction number) is the expected number of secondary cases directly generated by one case. Informally, the number of people each individual is likely to infect. Big R0 bad; R0 < 1 good.

And there are unfortunately the “super spreaders” - individuals who infect many people. [NYT (March 23, 2020)]

So far there's been lots of speculation for asymptomatic carrier rate

Mild or no symptoms: 60%

”Some degree” - Fauci

Mild or no symptoms: 86%

We really need to pin this down.

It really influences how we control the disease. Eric Topol

covid-19

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4. Molecular tests for detection

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4. Molecular tests for detection

https://covid19map.tech/

Some countries are testing much more than others. What does this do to our calculations for rates of infection and mortality? Also, for risk factors? And environmental effects?

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4. Molecular tests for detection

The standard test currently is based on measuring RNAs shed by the virus, and remaining in the throat.They are doing drive-in testing now in downtown Montreal.

Two steps:

  1. Sample Collection. Throat swab. This is the protocol that Ontario uses here. Very simple.
  2. Detect RNA form SARS-CoV-2 in the sample. RT-PCR assay for the detection.

Corman et al. (2019) Euro Surveill [from the WHO’s website here]

Not rocket science, but … recommended by SOP from WHO so comparable and guaranteed to work (doesn’t mean it is perfect; there is a whole field of biostatistics that address accuracy).

In the USA, the CDC sent out kits with a faulty control problem for the RT-PCR. This delayed testing [MIT-Press (March 5 2020)].

The CDC’s protocol requires the use of a specific Qiagen RNA extraction minikit.

But there is a big shortage of those kits.This is the first bottleneck in testing (for example, in the USA)!

This paper Bruce et al. (March 20, bioRxiv) looks at alternative extraction kits and tests not

based on RNA extraction!

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Diagnostic test for COVID-19 from Simon Fraser University, Vancouver

  • Based on a reagent prepared for high-resolution imaging of RNA molecules in vivo and fixed cells.
  • Mango II (after its bright color) binds to SARS-CoV-2 RNA and

fluorescences.

  • Research article is here, short presentation is here.

[Source: Simon Fraser University]

4. Faster, more performant molecular tests in the works

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4. Molecular tests for determining if you have had the disease

We recommend [Searchinger et al. (March 24 2020) Washington Post]

Serological tests are being developed to detect SARS-CoV-2 antibodies in people who suspect they might have

had Covid-19 (i.e. the ‘recovered’ pink people in the epidemiology simulation; slide 48).

[Amanat et al. (March 18, 2020) medRxiv]

Enzyme-linked ImmunoSorbent Assays (ELISA) - detects antigens in a patient’s serum (blood) derived from the spike (S) protein of SARS-CoV-2

Can be used to screen health care workers and minimize the risk of viral spread, if we assume that recovery means immunity

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South Korea just created a 10 minute antibody detection kit.

  • Uses immunochromatography
  • Detects COVID-19 IgM/IgG antibodies in blood, serum, or plasma produced in response to the infection
  • These antibodies represent an immune response to the virus
    • Useful for detecting infections that are asymptomatic
  • Shipping 3 million units this week.

Protocol of ELISA test with similar purpose made available by Krammer lab, Mount Sinai (@florian_krammer)

[PREVIOUS SLIDE]

4. Molecular tests for determining if you have had the disease.

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3. Epidemiology of SARS-Cov-2

The current approach to control the pandemic perhaps broadly based on two components:

non-pharmaceutical interventions + disease diagnostics

(eg social distancing) (eg throat swab-based RNA testing)

This “buys us MORE time” to develop

pharmaceutical interventions + herd immunity

(eg a vaccine)

Testing provides a type of “feedback” to optimize our interventions (red arrow) which buys us more time to improve our non-pharmaceutical interventions.

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4. Molecular tests for detection

“Skate to where the puck is going, not where it has been.”

  • Walter Gretzky, Canadian proverb

It is interesting to look at how some countries “contained” covid-19.

China: Lockdowns & electronic surveillance

[Kuperschmidt, Cohen (March 2 2020) Science]

[Cyranoski (March 27 2020) Nature]

South Korea: Through large-scale testing [Normile (March 17 2020) Science]

Privacy versus containing the disease without lockdown[Zastrow (March 18 2020) Nature]

Taiwan: How big data analytics were used [Wang et al. (March 3, 2020) JAMA]

This might give some indication of how we might proceed.

Some countries where it is not clear why they have had lower infection rates

Japan

Some countries do not seem to be testing at all (at least they weren’t testing earlier).

Russia

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5. Drugs and Therapeutics

One potential therapeutic strategy is based on chloroquine (CQ)

[Liu et al. (March 18, 2020) Cell Discovery]

Big in the news right now; 7 clinical trials underway Chinese Clinical Trial Registry for COVID-19.

  • CQ = N4-(7-Chloro-4-quinolinyl)-N1, N1-diethyl-1,4-pentanediamine
  • CQ used to treat malaria and amebiasis

  • hydroxychloroquine (HCQ) sulfate
  • weak bases
  • used already (that is, “approved”) for autoimmune disease incl. lupus and rheumatoid arthritis
  • immunomodulator
  • alkalinize/enlarge endo-lysosomes
  • trap internalized SARS-CoV-2
  • inhibit release of virus
  • combination therapy HCQ + azithroymycin (stimulates an immunomodulatory effect)
  • overblown? @jpogue1

Another potential therapeutic is Remdesivir, a nucleoside analog prodrug developed by Gilead Sciences Inc.

  • antiviral drug developed to combat Ebola; nucleotide analog
  • may have activity against various single stranded RNA viruses like SARS.
  • Not a lot available currently. Didn’t make it into production in 2003/4
  • phase III clinical trial (that is, “advanced”) launched in Wuhan,

February 4, 2020. We recommend

[Wang et al. (Feb 4 2020) Nature Cell Research]

[Kupferschmidt (March 22 2020) Science]

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5. Many new creative approaches for pharmaceutical interventions

This paper is one important step towards inhibiting the main protease of covid-19.

Specifically, they have been able to inhibit

the main protease with small molecules…

Next Slide Please

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Rapid science to understand the molecular structure of SAR-CoV-2

The main protease of SARS-CoV-2 is bound and blocked by a alpha-ketoamide inhibitor [Zhang et al. (March 20, 2020) Science]

One of the best characterized drug targets among coronaviruses. Inhibiting the activity of this enzyme would block viral replication. No human proteases have a similar cleavage specificity, so it likely wouldn’t be a toxic drug. This paper shows that this protease in SARS-CoV-2 is targettable.

5. Other Drugs and Therapeutics?

5. Many new creative approaches for pharmaceutical interventions

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5. Drugs and Therapeutics

There are actually many therapeutics in the “pipeline”. These and more ….

[The Economist]

Actemra (tocilizumab) is a monoclonal antibody against interleukin-6 approved for rheumatoid arthritis. It’s been approved for clinical trials on COVID-19 patients by the FDA.

We recommend this article about Roche’s Actemra drug that was recently approved.

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Mild to Moderate Cases

Drugs used in managing COVID-19 cases are experimental and the full extent of their action is still unknown (although they are approved for other diseases). The following protocol is used in patients tested positive for COVID19 using PCR:

  • Hydroxychloroquine or Chloroquine
  • If contraindicated; combination of Lopinavir and Ritonavir
  • Ribavirin in combination with Lopinavir/Ritonavir
  • Antipyretics in case of fever
  • Oxygen Therapy only if oxygen saturation (SpO2) below 93%
  • Supportive measures (fluids, electrolytes) and close monitoring

Treatment protocols for severe/critical cases are more aggressive and require ICU admittance along with mechanical ventilation.

Treatments are still experimental and little is known about their efficacy. Protocols are evolving rapidly.

For clinicians, SARS, and specifically SARS-CoV-2 here, are difficult to treat. Canadian doctors are currently learning as quickly as they can from regions of the world already affected by covid-19.

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6. Clinical Treatment

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Mechanism of action of these drugs:

Chloroquine/Hydroxychloroquine: Believed to limit endosomal acidification (virus requires acidic conditions to infect cells) and terminal glycosylation of ACE2 receptor. This leads to reduced receptor binding.

Lopinavir/ritonavir: Antiretroviral protease inhibitors. Results in immature and non-infectious viral particles and prevents infection of susceptible cells. Ritonavir also inhibits breakdown of lopinavir which consequently increases its half-life.

Ribavirin: Guanosine analogue, inhibits viral polymerases. Also, increases mutation rate in viruses, making them unfit (due to excessive mutations).

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6. Clinical Treatment

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Vaccines

We recommend the NEWS EXPLAINER available from [Callaway (March 18, 2020) Nature]

Many groups are working on vaccines; it’s likely to take a while. There are dangers of rushing, like vaccine enhancement, where a vaccine can make subsequent infections more rather than less severe [Jiang (March 16 2020) Nature]

Perhaps the best target for a vaccine against SARS-COV-2 is the spike protein on the surface of the virus.

There are still no approved vaccines against two previous Coronaviruses, SARS-CoV and MERS-CoV. Therefore there is uncertainty about the safety of any vaccine for SARS-CoV-2.

Decreasing levels of antibodies have been observed in patients infected by SARS-CoV and MERS-CoV. Meaning, the amount of antibody is insufficient to protect the person previously infected by these coronaviruses after 2 or 3 years.

That issue must be addressed, and taken into consideration in how the vaccine is administered, to prevent another pandemic in future years. Who should be given a vaccine (assuming there is enough): everyone? the elderly? people with underlying health conditions? health care workers? Does the answer depend on how long the vaccine remains effective? see answers for the Ebola vaccine.

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6. Clinical Treatment

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6. Clinical Treatment

Perhaps create new immunotherapeutic strategies

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7. Pandemic effects on air pollution

Concentrations of nitrogen dioxide (NO2) have decreased dramatically over Asian and European cities following decreases in travel and industrial activity. Nitrogen dioxide is not a greenhouse gas, but is a pollutant produced from car engines, power plants and other industrial processes that has serious impacts on ecosystems and human health. [The Guardian (March 23)]

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Pollution levels in China in 2019, left, and 2020. Photograph: Guardian Visuals / ESA satellite data

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7. Pandemic effects on global atmospheric CO2?

Ralph Keeling estimated that a sustained 10% drop in CO2 emissions would cause CO2 in the atmosphere to deviate by roughly 0.5 ppm.

#Keeling_Curve #CO2 #CO2emissions #coronavirus #COVID19

No events in the 62-year history of the Keeling Curve – including the downturn of 2008 and the Soviet Union collapse in late 1980s – have caused such a drop to date.

However, CO2 emissions from China have dropped by 25 % since the beginning of the outbreak. That represents a 6 % drop in global emissions.

History has shown that carbon dioxide levels typically resume their climb quickly as economic activity rebounds.

If there is any benefit of the SARS-CoV-2 event in terms of slowing the pace of climate change, it could be in changes in travel and economic activity that lead to sustained reductions in fossil fuel use.

Only those kinds of long-term systemic reductions will change the trajectory of CO2 levels in the atmosphere, Keeling said.

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Open Book Exam Question #3 [33 marks out of 100], 18 months to complete.

Meditate on any or all of the following concepts:

  • There are obviously many additional facets to a pandemic than those described here. For example, human psychology and social behavior. Where in the scientific literature would you look to explore these topics relative to covid-19?
  • How would moving too fast (i.e. reckless) towards a vaccine be counterproductive in the long term?
  • What is the role of science communication in a pandemic? What is the role of social media? How should scientists at all different levels of training and experience convey their knowledge?
  • How can we as scientists best contribute on an individual level and collectively to ensure we do far more good than harm?
  • How can we lead by example?

We recommend this ASD report covering the effects of covid-19 on different economic sectors. They have made it free of charge.

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Acknowledgements

C Gamberi M Whiteway

A Piekny

P Peres-Neto

M Hallett E Despland

Samira Massahi C Ziter

Aki Eftyhios Kirbizakis V Martin

Van Bettauer

Abdelrahman Ahmed

Vanessa Dumeaux, PhD Epidemiology

We thank Dr Thomas Nolan, MD, Respirologist for comments, insights and references.

We thank Prof. WD Lubell, medicinal chemist at UdeM, for comments and insights.

In the meantime, why not learn some bioinformatics?

Good starting point @lizilla93

Questions 1, 2, 3 sum to 99 of 100. If you stay healthy, +1.